1. Field of the Invention
The present invention relates to an apparatus for performing a continuous free flow electrophoresis process. More particularly, the present invention relates to an apparatus for performing a continuous free flow electrophoresis procedure which is constructed such that there are no ion permeable membranes between the electrode chambers and the central separation chamber.
2. Description of the Prior Art
Electrophoresis, in general, is the phenomenon of the migration of charged particles or ions in a liquid carrier medium under the influence of an electric field. This phenomenon can be used to separate small particles which, by reason of different surface chemical properties, exhibit different concentrations of surface charge in the given medium. Under the influence of the electrical field, the electrophoretic mobilities of the various classes of charged particles in the carrier medium will be different. A sample continuously introduced at some point into the sheet of liquid carrier medium (buffer) flows in a narrow band in the absence of a potential gradient; however, when the potential gradient is applied to the sheet of buffer, the sample particles are separated under the influence of the electrical field into various particle groups or components depending upon the electrophoretic mobility of the respective particles, the strength of the field, and the length of time that the particles remain in the field. Particles of similar mobility are concentrated in distinctive zones or bands which fan out from the point of sample introduction.
The present invention relates in particular to free flow continous electrophoresis in which a buffer solution is made to flow freely in a uniform film or sheet through a chamber defined by two parallel enlongate plates. A sample is introduced into the buffer sheet at some point, and an electric potential gradient is applied across this flowing sheet perpendicular to the direction of buffer flow. The individual components within each sample then separate into narrow bands, depending upon their respective electrophoretic mobilities, and can be collected from the outlet end of the electrophoresis chamber through one or more of a plurality of small tubes disposed along a collection manifold at the outlet of the chamber.
In the present art, the electrical field is established between a pair of electrodes, one at each side of the separation chamber. Because the carrier fluid or buffer is normally water plus ionic species added to protect the viability of the biological material being separated, the buffer fluid is electrically conductive. When the electrical field is imposed upon the conductive buffer carrier fluid, electrolysis of the water occurs, liberating hydrogen gas at the cathode electrode and oxygen gas at the anode electrode. Since the amount of gas liberated usually exceeds the amount that can be dissolved in the carrier buffer flow, undesirable by-products are introduced into the buffer flow stream. These undesirable by-products cause instability affecting separation and should not be allowed to contaminate the buffer flow. To this end, conventional free flow electrophoresis separation apparatus are universally constructed with ion permeable membranes or ion exchange membranes separating the electrode chambers from the central separation chamber. The use of such membranes by no means renders the apparatus ineffective; however, the use of membranes does place substantial limitations upon the separation process. First, polarization of the membranes under the influence of the electric field causes variations in the population densities of the various ionic species contained within the separation chamber. Also, heating of the carrier (buffer) fluid is increased due to the resistance of the ion permeable membranes to the flow of electrical current through them. Further, pressure differentials across the ion permeable membranes are introduced by the mechanism of electro-osmotic pumping.